RESEARCH PROJECT 2 ? SUMMARY Synovial sarcoma (SS) is a paradigm of a gene fusion driven cancer, in which the defining event is the translocation t(X,18; p11, q11), that creates an in-frame fusion of the SS18 gene to SSX1, SSX2 or SSX4 genes. Conventional therapies for SS, including cytotoxic drugs (doxorubicin, ifosfamide), provide limited benefit, leaving patients at risk for local recurrence and metastases. Despite best available therapies, this high-grade sarcoma leads to death in approximately 25% of patients within 5 years. Unlike other soft tissues sarcomas translocations in which a transcription factor is thought to confer target specificity by binding DNA in a sequence specific manner, SS18-SSX lacks a DNA binding domain and apparently exerts its activity by interacting with other chromatin regulators. We recently identified a critical role for KDM2B and other members of the non-canonical polycomb repressive complex 1 (PRC1.1) in sustaining SS cell proliferation by mediating SS18-SSX chromatin occupancy. Here we propose to develop embryonic-stem cell (ESC)-based genetically engineered mouse models to evaluate the therapeutic efficacy of inhibiting SS18-SSX, PRC1.1 or new targets required for SS18-SSX activity and assess the potential toxicities of such treatments in fully developed mice. This new modelling strategy will dramatically increase the speed and scale at which potential therapeutic targets can be investigated for synovial sarcoma. Additionally we propose to develop strategies for targeting SS18-SSX oncogenic activity by combining an array of biochemical analysis, functional genomics and genetically engineered mouse models. We aim to identify SS18-SSX protein domains as well as protein interaction networks to develop methods to inhibit its chromatin localization. Having recently characterized SS18-SSX genome-wide chromatin occupancy and direct gene targets, we will use functional genomics to dissect pathways operating downstream of this key oncogenic driver. Further, given the presence of mutations in the H3K36me3 methylatransferase - SETD2 - in 5-10% of synovial sarcomas we will define the role of H3K36 methylation in synovial sarcoma by taking advantage of patient data, functional assays and mouse models generated in this proposal.